The major goal of this research project is to elucidate the function of the small GTPase Rapl and the molecular mechanism(s) by which this GYPase exerts its effects on epithelial morphogenesis. Recent studies in Drosophila and in mammalian systems have implicated Rapl proteins in the control of cell spreading, adhesion and morphogenesis, but little is known about the molecular mechanisms that mediate Rapl function. The mammalian junctional protein AF-6 was identified as a Rapl-interacting protein, but functional analyses of mammalian Rap 1 and AF-6 proteins, and of their interaction, have been hampered by the fact that these proteins are largely refractory to loss- and/or gain-of-function analysis. Hence, we decided to first gain insights into their functions using a genetically more tractable Drosophila system. Our preliminary results from Drosophila implicate Canoe (the Drosophila homologue of AF-6) as an effector of DRapl (the Drosophila homologue of Rapl) in dorsal closure (DC), a morphogenetic process that relies on elongation and migration of epithelial sheets in the embryo. Furthermore, we have recently identified a putative novel regulator of DRapl in DC, designated DPDZ-GEF, whose vertebrate homologue is associated with epithelial adherens junctions, just as Rapland AF-6/Canoe are. Notably, the Drosophila homologues of Rapl and AF-6 (DRapl and Canoe) share a similar physical interaction, and the mammalian proteins can functionally substitute for their Drosophila counterparts. This proposal aims to define the function of DRapl, Canoe, and DPDZ-GEF in regulating cell shape changes, cell-cell adhesion, and coordinated movement of an epithelial sheet during DC. Our specific aims are: 1) To analyze the function of DRapl and Canoe in DC and cell-cell adhesion, and 2) To characterize the function of DPDZ-GEF. Specific aim 1 is designed to provide a thorough assessment of the mechanisms that underlie the DRapl/Canoe interaction and the role of Canoe as a junctional protein in the process of DC. In aim 2, the biochemical and biological functions of the DPDZ-GEF and its epistatic relationship to DRapl will be assessed. These studies offer a comprehensive set of approaches to define the roles of DRapl, Canoe, and DPDZ-GEF in regulating cell-cell adhesion and coordinated epithelial cell movements, and will likely lead to a better mechanistic understanding of these processes and facilitate the design of experiments addressing the role of the mammalian homologues in similar processes. Ultimately, our studies are relevant to a number of medically important issues, including wound healing and the homeostasis of epithelial cells and sheets.